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Title: Study of the endoplasmic reticulum proteostasis network during ageing
Author: Mares, Alina
Awarding Body: University of Manchester
Current Institution: University of Manchester
Date of Award: 2013
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The functional integrity of the proteome is essential for proper cell functioning. Protein homeostasis, or proteostasis, is maintained by a network of pathways that mediate the biosynthesis, folding and degradation of proteins. Accumulating evidence suggest that ageing is associated with a general decline in protein homeostasis. Proteins are synthesised in the cytosol as extended polypeptide chains, which must then be folded in to their native conformation, before moving to their site of function. Protein folding occurs in different subcellular compartments, such as the cytosol and the endoplasmic reticulum (ER). Due to the nature of the proteins that are processed in the ER (e.g. ionic channels, receptors, hormones, signalling molecules), the function of this compartment can be seen to be of vital importance to the cells. However, the effect of age on ER protein homeostasis is virtually unknown. A combination of post-mortem mouse tissues and cell-based models were used to examine the impact of age on ER protein folding, quality control and the ER stress response. Analysis of the expression level of ER-resident and ER-linked proteins showed a number of age-related changes in mouse tissues, in human fibroblasts aged in vitro by serial passage, and human fibroblasts obtained from young and old donors (in vivo aged cells). Overall, the pattern of changes was variable between different tissues and cell systems. However, a common feature of aged tissues and both cellular models of ageing, was a significant increase in phosphorylation of eIF2alpha, indicating that ER protein homeostasis is affected with age. In addition, ageing in several tissues and in both cellular systems was associated with accumulation of polyubiquitinated substrates, suggesting that degradation of abnormal proteins via the proteasome is deficient. More detailed investigation of ER proteostasis using the cellular models showed that in vitro aged cells had a decreased capacity to fold a temperature sensitive model membrane protein (ts O45 VGV-G) and were less efficient at degrading two model substrates of the ER-associated degradation pathway. Since the ER protein folding and degradation pathways are modulated by ER stress response signalling to restore ER homeostasis, therefore their malfunctioning would impact on the ability of aged cells to cope with stress. Indeed, in the aged cells both PERK and IRE-1 signalling were perturbed. In addition, the level of BiP was not upregulated following ER stress in the aged cells. Moreover, ER stress induction led to decreased cell survival in the case of aged cells, showing that the inability of aged cells to maintain ER protein homeostasis results in increased susceptibility to cell death. All these data together shows how perturbed ER proteostasis can occur with increased age and its impact on overall cell wellbeing, therefore provide new insight into mechanisms of cell ageing.
Supervisor: Boot-Handford, Raymond ; Swanton, Eileithyia Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: ageing ; ER proteostsis